Development of a Real-Time Polymerase Chain Reaction Detection Method for the Quantitation of Myeloperoxidase Transcripts in Porcine Tissues

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2001-01-01
Authors
Fitzsimmons, C.
Marklund, L.
Tuggle, C.
Stabel, T.
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Myeloperoxidase (MPO) is a hemoprotein present in azurophilic granules of polymorphonuclear (PMN) leukocytes and monocytes. It catalyzes the oxidation of halide ions to their respective hypohalous acids, which are used for microbial killing by phagocytic cells. Measurement of MPO activity is often used as a marker of neutrophil infiltration into tissues. We have designed a quantitative reverse transcription-polymerase chain reaction detection method for porcine MPO transcripts by using TaqMan realtime PCR technology. Total RNA was isolated from lung and spleen tissue collected 7 days post-intranasal inoculation with Salmonella choleraesuis (n=4) or saline (n=4). The lung and spleen samples were pooled before RNA isolation to create uninfected control and infected RNAs for each tissue. Expression of MPO mRNA was highest in infected spleen (12.5 ± 2.85 relative units (RU)), followed by the uninfected spleen sample (2.9 ± 1.50 RU). There was no difference in MPO expression between uninfected and infected lung samples. In conclusion, levels of MPO mRNA expression in porcine spleen and lung indicate a differential response to infection between the two tissues. This difference may be associated with bacterialhost adaptation of S. choleraesuis. The TaqMan assay for MPO also can be used to discover tissue-specific responses between individuals or groups of pigs exhibiting distinct phenotypic responses to infection.

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